A genetically engineered cell-based system for detecting metabolism-mediated toxicity

Xenobiotics undergong bioactivation by CYP450 enzymes form reactive metabol ites that may exert direct metabolism-mediated toxicity. An in vitro model was developed to study the direct toxic effects that follow the metabolic a ctivation of chemicals. The model uses monolayer cultures of genetically en gineered NIH-3T3 or V79 cells that express individual human or rat CYP450 i soforms, respectively. Following exposure to 1,3-dichloropropanol or cyclop hosphamide, basal cytotoxicity endpoints, including neutral red uptake and Alamar Blue(TM) reduction were used to assess changes in cell number and fu nctional viability resulting from the formation of metabolites. Cell lines that express cytochrome P450 enzymes metabolised the test compounds, leadin g to increased toxicity compared with that observed in the control cell lin e. The use of specific inhibitors confirmed that the formation of reactive metabolites was CYP450-isoform dependent. These results indicate that a pan el of genetically engineered cell lines expressing various cytochrome P450 enzyme isoforms can be used to reveal measurable metabolising capabilities, and could become a useful tool for the detection and possible determinatio n of CYP450 isoforms in human liver metabolism-mediated toxicity.